Rape oil, oxidized

Biodegradation in water: screening tests

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

biodegradation in water: screening test, other

Remarks:

According to CEC L-33-T-82

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

January - March 1989

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: According to GLP, but performed according to a guideline which measures only primary biodegradation; the concentration of the inoculum and the test material in the test flasks were not reported.

Qualifier:

according to guideline

Guideline:

other: CEC L-33-T-82

Deviations:

no

Principles of method if other than guideline:

The test determined degradation by measuring the disappearance of CH3-CH2-groups, typical for hydrocarbons. The flasks with test substance and inocolum were incubated for 21 days in the dark at 25°C for 21 days (no stirring or shaking). After 0, 7 and 21 days, the solutions were extracted with carbontetrachloride. The organic extracts were analysed using IR-spectroscopy at 2930 cm-1 (absorption band for CH3-CH2-groups).

GLP compliance:

yes

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic, non-adapted

Details on inoculum:

- Source of inoculum/activated sludge (e.g. location, sampling depth, contamination history, procedure): Domestic sewage treatment plant of Schmallenberg, Germany
- Laboratory culture: no
- Method of cultivation: not relevant
- Storage conditions: no data
- Storage length: no data
- Preparation of inoculum for exposure: centrifugation followed by CFU count by plate method.
- Pretreatment: none
- Concentration of sludge: no data
- Initial cell/biomass concentration: no data (CFU count of inoculum was 9*10^6 cells/mL).
- Water filtered: no
- Type and size of filter used, if any: not relevant

Duration of test (contact time):

21 d

Initial conc.:

other: no data

Parameter followed for biodegradation estimation:

other: IR-spectroscopy at 2930 cm-1 (absorption band for CH3-CH2-groups).

Details on study design:

Triplicate Erlenmeyer flasks (size not reported, closed with cottonwool stoppers allowing gaseous exchange) with test substance and inocolum (volume of incubate and concentrations of test substance and inoculum not reported) were incubated for 21 days in the dark at 25°C for up to 21 days (no stirring or shaking). Duplicate toxic control flasks with HgCl2 and inocolum (concentrations of HgCl2 and inoculum not reported) were incubated in the same way. After 0, 7 and 21 days, the test solutions were extracted with carbontetrachloride. The organic extracts were analysed using IR-spectroscopy at 2930 cm-1 (absorption band for CH3-CH2-groups). Two calibration controls containing test substance dissolved in carbontetrachloride were measured by IR-spectroscopy on day 0 to serve as initial 100% value. Neutral controls (inocolum but no test substance) were included and the absorbance of the neutral controls at day 0 was subtracted from that of the test flasks. Toxic control flasks were sampled and analysed as described above after 7 and 21 days. The data obtained for the toxic control were used to correct for degradation due to other than biological processes.

Reference substance:

other: di-iso-tridecyladipate

Parameter:

other: Disappearance of CH2-CH2-groups

Value:

60.4

Sampling time:

7 d

Parameter:

other: Disappearance of CH2-CH2-groups

Value:

81.1

Sampling time:

21 d

Details on results:

see below

Results with reference substance:

35.9% degradation after 7 days, 90.4% degradation after 21 days.

A summary of the results is provided below.

Day 0.

In triplicate test flasks, the mean extinction on day 0 (corrected for the extinction in neutral flasks) was 0.573 (CV 0.8%). The extinction of the calibration solutions was 0.580, hence the recovery was 98.9%. Since the CV was <5% and the recovery was between 95% and 105%, the test was considered valid.

Day 7.

In triplicate test flasks, the mean extinction on day 7 (corrected for the extinction in neutral flasks) was 0.201 (individual values 0.079, 0.263, 0.262). In duplicate toxic flasks, the mean extinction on day 7 (corrected for the extinction in neutral flasks) was 0.509 (individual values 0.508, 0.509). The amount of test substance in the test flasks and the toxic flasks was 35.1% and 88.7%, respectively, of the initial value and the percentage degradation of the test substance corrected for abiotic degradation was 60.4%.

Day 21.

In triplicate test flasks, the mean extinction on day 7 (corrected for the extinction in neutral flasks) was 0.096 (individual values 0.115, 0.056, 0.116). In duplicate toxic flasks, the mean extinction on day 7 (corrected for the extinction in neutral flasks) was 0.506 (individual values 0.501, 0.510). The amount of test substance in thetest flasks and the toxic flasks was 16.7% and 88.2%, respectively, of the initial value and the percentage degradation of the test substance corrected for abiotic degradation was 81.1%.

Validity criteria fulfilled:

yes

Interpretation of results:

other: 60.4% and 81.1% primary degradation after 7 and 21 days, respectively

Conclusions:

In a test according to CEC L-33-T-82, 60.4% and 81.1% primary degradation of rapeseed oil REM 8/60 P was recorded after 7 and 21 days, respectively. The interpretation of the result is hampered by the lack of information on the test substance and the inoculums concentration in the test flasks.

Reference 2

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

February 10, 1988 - March 31, 1988

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Remarks:

According to GLP, but not according to a recommended OECD 301 or 302 guideline, and with reporting deficiencies.

Qualifier:

according to guideline

Guideline:

other: Method for the determination of ecotoxicity at level 1, Biodegradation; Repetitive Die Away test. DG XI/400/84. Rev.1, EEC, 1984

Deviations:

no

Remarks:

insufficient information to evaluate compliance

Principles of method if other than guideline:

The test medium is inoculated with activated sludge, stabilized for one week and then spiked with the test substance. After two weeks, three repetitive weekly additions of test substance take place. After each repetitive addition, the water phase is re-aerated to the saturation level. The oxygen consumption is measured weekly up to two weeks after the last addition. The percentage biodegradation is calculated from the measured oxygen consumption and the analytically determined Chemical Oxygen demand (COD).

GLP compliance:

yes

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic, non-adapted

Details on inoculum:

- Source of inoculum/activated sludge (e.g. location, sampling depth, contamination history, procedure): Municipal waste water treatment plant of Duiven, The Netherlands, receiving only waste water from domestic origin
- Laboratory culture: no
- Method of cultivation: not relevant
- Storage conditions: no data
- Storage length: preconditioned during a week (unspecified conditions)
- Preparation of inoculum for exposure: no data
- Pretreatment: no data
- Concentration of sludge: final concentration in test flasks 35 mg suspended solids/L.
- Initial cell/biomass concentration: no data
- Water filtered: no data
- Type and size of filter used, if any: no data

Duration of test (contact time):

14 d

Initial conc.:

40 mg/L

Based on:

test mat.

Initial conc.:

ca. 120 mg/L

Based on:

ThOD/L

Parameter followed for biodegradation estimation:

O2 consumption

Details on study design:

The test medium was inoculated with activated sludge (35 mg suspended solids/mL), stabilized for one week and then spiked with the test substance (40 mg/L). The following emulsifiers were used: genapol PF-40, 10% of the weight of the oil; Nonylphenol.10EO.5PO, 20% of the weight of the oil. It was reported that these emulsifiers are not toxic in the biodegradation test and not biodegradable. Control solutions contained inoculums and emulsifier but no test substance. Triplicate flasks (dark glass bottles containing 187 mL of liquid and 93 L of gas space) were used for the treated and the control test solutions. The flasks were incubated at 20°C and were shaken continuously to assure steady state oxygen partitioning between the liquid and the gas phase. After two weeks, three repetitive weekly additions of test substance were performed. After each repetitive addition, the water phase was re-aerated to the saturation level. The oxygen consumption was measured weekly up to two weeks after the last addition. The percentage biodegradation was calculated from the measured oxygen consumption in the treated test solutions, corrected for the oxygen consumption in the control solutions, and the analytically determined Chemical Oxygen Demand (2.07 g/g) of the test substance. A toxicity control was included (sodium acetate in the presence of 7.6 mg oil/L).

Reference substance:

acetic acid, sodium salt

Remarks:

concentration not reported

Parameter:

% degradation (O2 consumption)

Value:

38

Sampling time:

7 d

Parameter:

% degradation (O2 consumption)

Value:

45

Sampling time:

14 d

Parameter:

% degradation (O2 consumption)

Value:

30

Sampling time:

21 d

Remarks on result:

other: total % biodegradation for first two additions one week after second addition

Parameter:

% degradation (O2 consumption)

Value:

29

Sampling time:

28

Remarks on result:

other: total % biodegradation for first three additions one week after third addition

Parameter:

% degradation (O2 consumption)

Value:

35

Sampling time:

42

Remarks on result:

other: total % biodegradation for four additions two weeks after fourth addition

Details on results:

see below

Results with reference substance:

About 70% biodegradation within one week.

The results are summarised in the table below. These results were derived from mean values for oxygen concentrations in test solutions. The individual replicate values were not reported.

 

Table 1. Results of Repetitive Die Away test with blown rapeseed oil, 60 P.

parameter	day 0	day 7	day 14	day 21	day 28	day 35	day 42
total oxygen uptake (mg)	-	5.9	7.0	9.5	13.8	18.4	21.7
total oxygen demand (mg)	15.7	15.7	15.7	31.4	47.1	62.8	62.8
% biodegradation	-	38	45	30	29	29	35

 

Validity criteria fulfilled:

not specified

Interpretation of results:

inherently biodegradable, not fulfilling specific criteria

Conclusions:

In a test measuring oxygen uptake (not according to any OECD 301 or 302 guideline), 45% biodegradation was recorded after 14 days exposure to non-adapted inoculums, and 35% biodegradation two weeks after the last of three subsequent weekly additions to adapted inoculum. The rate of biodegradation may have been impaired by problems of solubility and mass transfer.

Reference 3

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

key study

Study period:

27-jan-2010 to 25-feb-2010

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

This study has been performed according to OECD and/or EC guidelines and according to GLP principles

Qualifier:

according to guideline

Guideline:

OECD Guideline 301 B (Ready Biodegradability: CO2 Evolution Test)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.4-C (Determination of the "Ready" Biodegradability - Carbon Dioxide Evolution Test)

Deviations:

no

Qualifier:

according to guideline

Guideline:

ISO DIS 9439 (Ultimate Aerobic Biodegradability - Method by Analysis of Released Carbon Dioxide)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic, non-adapted

Details on inoculum:

The source of test organisms was activated sludge freshly obtained from a municipal sewage treatment plant: 'Waterschap de Maaskant', 's-Hertogenbosch, The Netherlands, receiving predominantly domestic sewage.

Treatment : The freshly obtained sludge was kept under continuous aeration until further treatment. The concentration of suspended solids was 3.8 g/l in the concentrated sludge (information obtained from the municipal sewage treatment plant). Before use, the sludge was allowed to settle (73 minutes) and the liquid was decanted for use as inoculum at the amount of 10 ml/l of mineral medium.

Duration of test (contact time):

28 d

Initial conc.:

32 mg/L

Based on:

test mat.

Initial conc.:

12 mg/L

Based on:

other: Total Organic Carbon (TOC)

Parameter followed for biodegradation estimation:

CO2 evolution

Details on study design:

TEST CONDITIONS
- Composition of medium: 1 litre mineral medium contains: 10 ml of solution (A), 1 ml of solutions (B) to (D) and Milli-RO water
Stock solutions of mineral components
A) 8.50 g KH2PO4; 21.75 g K2HPO4; 67.20 g Na2HPO4.12H2O; 0.50 g NH4Cl; dissolved in Milli-Q water and made up to 1 litre, pH 7.4 ± 0.2
B) 22.50 g MgSO4.7H2O dissolved in Milli-Q water and made up to 1 litre.
C) 36.40 g CaCl2.2H2O dissolved in Milli-Q water and made up to 1 litre.
D) 0.25 g FeCl3.6H2O dissolved in Milli-Q water and made up to 1 litre.

- Test temperature: between 21.7 and 22.2°C.
- pH:
At t=0 d: 7.5 - 7.6
At t=28 d: 7.7 - 8.0
- pH adjusted:no
- Aeration of dilution water: Not before the test, the test is aerated continously
- Suspended solids concentration: The concentration of suspended solids was 3.8 g/l in the concentrated sludge (information obtained from the municipal sewage treatment plant).
- Continuous darkness: yes

TEST SYSTEM
- Culturing apparatus: 2 litre all-glass brown coloured bottles
- Number of culture flasks/concentration:
Test suspension: containing test substance and inoculum (2 bottles).
Inoculum blank: containing only inoculum (2 bottles)
Positive control: containing reference substance and inoculum (1 bottle).
Toxicity control: containing test substance, reference substance and inoculum (1 bottle).
- Method used to create aerobic conditions:
Synthetic air (a mixture of oxygen (ca. 20%) and nitrogen (ca. 80%)) was sparged through the solutions at a rate of approximately 1-2 bubbles per second (ca. 30-100 ml/min).
- Test performed in open system: yes
- Details of trap for CO2 and volatile organics if used:
CO2 was trapped in barium hydroxide solution. The amount of CO2 produced was determined by titrating the remaining Ba(OH)2 with 0.05 M standardized HCl (1:20 dilution from 1 M HCl (Titrisol® ampul). Titrations were made every second or third day during the first 10 days, and thereafter at least every fifth day until the 28th day, for the inoculum blank and test suspension. Titrations for the positive and toxicity control were made at least 14 days.


SAMPLING
- Sampling frequency: Titration were made on day: 2, 5, 7, 9, 14, 19, 23, 27 and 29
- Sampling method: Titration of the whole volume of CO2-absorber

CONTROL AND BLANK SYSTEM
- Inoculum blank: yes
- Abiotic sterile control: no
- Toxicity control: yes
- Other:

Reference substance:

acetic acid, sodium salt

Parameter:

% degradation (CO2 evolution)

Value:

19

Sampling time:

29 d

Remarks on result:

other: HCl added on the 28th day (last CO2-measurement on the 29th day)

Parameter:

% degradation (CO2 evolution)

Value:

38

Sampling time:

29 d

Remarks on result:

other: HCl added on the 28th day (last CO2-measurement on the 29th day).

Details on results:

The relative biodegradation values calculated from the measurements performed during the test period revealed 19 and 38% biodegradation of Blown rapeseed oil, for the duplicate bottles tested. Thus, the criterion for ready biodegradability (at least 60% biodegradation within a
10-day window) was not met.

In the toxicity control more than 25% biodegradation occurred within 14 days (40%, based on ThCO2). Therefore, the test substance was assumed not to inhibit microbial activity.

Results with reference substance:

The positive control substance was biodegraded by at least 60% (77%) within 14 days.

Validity criteria fulfilled:

yes

Interpretation of results:

other: Not readily biodegradable under the conditions of the OECD Guideline, without any additional improved bioavailability conditions except agitation with this water insoluble substance.

Conclusions:

Not readily biodegradable under the conditions of the OECD Guideline, without any additional improved bioavailability conditions except agitation with this water insoluble substance.

Reference 4

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

key study

Study period:

December 2008 - May 2010

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

Conducted in accordance with OECD 301F, but not GLP

Qualifier:

according to guideline

Guideline:

OECD Guideline 301 F (Ready Biodegradability: Manometric Respirometry Test)

GLP compliance:

no

Remarks:

Study was not performed under GLP

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic, non-adapted

Details on inoculum:

- Source of inoculum/activated sludge: Municipal STP
- Storage length: 24 hours maximum
- Concentration of sludge: 30 mg/L SS

Duration of test (contact time):

28 d

Initial conc.:

100 mg/L

Based on:

COD

Initial conc.:

40 mg/L

Based on:

test mat.

Parameter followed for biodegradation estimation:

O2 consumption

Details on study design:

TEST CONDITIONS
- Composition of medium: in accordance with OECD Guideline
- pH:7.4 +/- 0.2
- pH adjusted: yes
- Suspended solids concentration: 30 mg/L
- Continuous darkness: yes

TEST SYSTEM
- Culturing apparatus: OxiTop - Control - testsystem
- Number of culture flasks/concentration: 2 (for each batch); 16 replicates in total

SAMPLING
- Sampling frequency:
- Sampling method:
- Sterility check if applicable:
- Sample storage before analysis:
- Other:

CONTROL AND BLANK SYSTEM
- Inoculum blank: yes
- Abiotic sterile control: no
- Toxicity control: no

Reference substance:

ethylene glycol

Parameter:

% degradation (O2 consumption)

Value:

> 10

Sampling time:

2 d

Remarks on result:

other: For the replicates

Parameter:

% degradation (O2 consumption)

Value:

>= 57.8 - <= 78.3

Sampling time:

10 d

Remarks on result:

other: Range of the averages of the 2 replicates for the 8 batches studied

Parameter:

% degradation (O2 consumption)

Value:

> 60

Sampling time:

12 d

Remarks on result:

other: For the replicates

Parameter:

% degradation (O2 consumption)

Value:

>= 74.1 - <= 99.6

Sampling time:

28 d

Remarks on result:

other: Range of the averages of the 2 replicates for the 8 batches studied

Details on results:

The data have been corrected for the inoculum blank in accordance wtih the OECD Guideline (checked from raw data separately reported). For all replicates, the raw data are available, showing that the 10-day window has been met in all the replicates: a 10% biodegration was reached after 2 days and after 12 days the biodegradation was >60%

Validity criteria fulfilled:

yes

Remarks:

Oxygen uptake of the inoculum blank is 15.5 mg/L in 28 days.

Interpretation of results:

readily biodegradable

Conclusions:

Readily biodegradable

Description of key information

Readily biodegradable (OECD 301F)

Key value for chemical safety assessment

Biodegradation in water:

readily biodegradable

Additional information

In the OECD 301B study, the criterion for ready biodegradability (at least 60% biodegradation within a 10-day window) was not met (19 and 38% biodegradation for the 2 duplicate bottles tested). On the other hand, in total 16 replicates have been studied in the OECD 301F study. In all these studies the criterion for ready biodegradability was met. Therefore it is considered justified to conclude that the substance is readily biodegradable. In the OECD 301B study, the rate of biodegradation may have been impaired by problems of solubility and mass transfer.